PTHrP was discovered as the tumor product that mediates most instances of humoral hypercalcemia. The PTHrP and PTH genes are related on the basis of an ancient duplication event and a consequence of this heritage is a highly homologous sequence at the N-terminus of both peptides and the fact that both N-terminal products seem to signal via the same receptor, the type I PTH/PTHrP receptor. PTHrP and the type I receptor are expressed in adult and fetal tissues in a hand-in-glove fashion suggesting a paracrine function. Recent experiments in mice have provided compelling evidence that one such PTHrP function is a developmental regulatory molecule. In the body of this proposal, we show that the ablation of PTHrP expression in the microenvironment of the tooth results in a failure to form an eruption pathway and that the transgenic replacement of PTHrP in the enamel epithelium restores the normal program of tooth eruption. We propose to investigate the mechanism by which PThrP regulates the process of tooth eruption in the following manner. Aim 1 involves the characterization of PTHrP function in vivo by a) histological comparison of the impaction process in rescued PTHrP- knockout animals relative to that in osteopetrotic src-null mice, b) a temporal-spatial developmental survey by in situ hybridization of the normal expression pattern of PTHrP and the PTH/PTHrP receptor in the mouse molar and incisor, c) analysis of the number, distribution and differentiation of osteoclasts in the alveolar bone of recused PTHrP- knockout mice, d) histomorphometric analysis of the alveolar bone in rescued PTHrP-knockout mice as an indicator of osteoclast function, and e) histological analysis of the impaction process in PTH/PTHrP receptor/knockout mice. Aim 2 proposes the establishment of primary culture systems for a) rodent dental epithelium and mesenchyme, and b) rodent osteoblasts, in order to directly test the production of PTHrP-treated dental follicle cells of a factor which stimulates the activity of osteoclasts. Osteoclast activity will be measured in a functional assay of bone resorption. Aim 3 proposes the identification of the presumptive resorptive factor by a) characterization of its basic biochemical properties, b) testing for the expression of known bone-resorbing factors by PTHrP-treated dental follicle cells, and c) the use of a PCR-based subtractive hybridization method to isolate mRNAs which are differentially expressed in PTHrP-treated dental follicle cells.